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<div id="Sx1" class="ltx_section">
<h1 class="ltx_title ltx_title_section">Camera setup</h1>

<div id="Sx1.p1" class="ltx_para">
<p class="ltx_p">As ThunderSTORM uses real units (photons, nanometers), accurate camera
parameters need to be supplied by the user. The effective pixel size
can be determined from the actual pixel pitch of the camera sensor
and from the total magnification of the optical setup, or by performing
a spatial calibration using a stage micrometer. The detector response
(photoelectrons per A/D count and base level offset of the camera
digitizer) can be obtained from the specification sheet supplied by
the camera manufacturer or from photon transfer curve measurements
(the dependence of the variance on the mean value of pixel intensities),
see e.g., <cite class="ltx_cite">[<a href="#bib.bib27" title="CCD charge collection efficiency and the photon transfer technique" class="ltx_ref">1</a>, <a href="#bib.bib37" title="Image detectors for digital image microscopy" class="ltx_ref">2</a>]</cite>. Correct pixel size
is important for proper spatial calibration of the rendered images.
Camera conversion gain and offset influence simulated images and the
estimates of localization precision.</p>
</div>
<div id="Sx1.SSx1" class="ltx_subsection">
<h2 class="ltx_title ltx_title_subsection">Pixel size </h2>

<div id="Sx1.SSx1.p1" class="ltx_para">
<p class="ltx_p">Pixel size is used to convert molecular coordinates from pixels to
nm. It is the size of one camera pixel as projected to the sample
plane.</p>
</div>
</div>
<div id="Sx1.SSx2" class="ltx_subsection">
<h2 class="ltx_title ltx_title_subsection">Conversion of image intensity to photoelectrons</h2>

<div id="Sx1.SSx2.p1" class="ltx_para">
<p class="ltx_p">For CCD cameras, the image intensity is converted to the number of
photoelectrons according to the formula</p>
<table id="Sx1.Ex1" class="ltx_equation">

<tr class="ltx_equation ltx_align_baseline">
<td class="ltx_eqn_pad"></td>
<td class="ltx_align_center"><img id="Sx1.Ex1.m1" class="ltx_Math" style="vertical-align:-6px" src="mi/mi2.png" width="108" height="21" alt="i=p\left(I-b\right)\,,"></td>
<td class="ltx_eqn_pad"></td>
</tr>
</table>
<p class="ltx_p">and for EMCCD cameras, the conversion is according to the formula</p>
<table id="Sx1.Ex2" class="ltx_equation">

<tr class="ltx_equation ltx_align_baseline">
<td class="ltx_eqn_pad"></td>
<td class="ltx_align_center"><img id="Sx1.Ex2.m1" class="ltx_Math" style="vertical-align:-18px" src="mi/mi1.png" width="112" height="39" alt="i=\frac{p}{g}\left(I-b\right)\,."></td>
<td class="ltx_eqn_pad"></td>
</tr>
</table>
<p class="ltx_p">Here <img id="Sx1.SSx2.p1.m1" class="ltx_Math" style="vertical-align:-2px" src="mi/mi6.png" width="10" height="16" alt="i"> is the number of photolectrons, <img id="Sx1.SSx2.p1.m2" class="ltx_Math" style="vertical-align:-5px" src="mi/mi7.png" width="13" height="15" alt="p"> is the CCD sensitivity
(in photons per A/D count), <img id="Sx1.SSx2.p1.m3" class="ltx_Math" style="vertical-align:-5px" src="mi/mi5.png" width="13" height="15" alt="g"> is the EM gain for EMCCD cameras,
<img id="Sx1.SSx2.p1.m4" class="ltx_Math" style="vertical-align:-2px" src="mi/mi3.png" width="13" height="16" alt="I"> is the image intensity (in A/D counts), and <img id="Sx1.SSx2.p1.m5" class="ltx_Math" style="vertical-align:-2px" src="mi/mi4.png" width="12" height="16" alt="b"> is the base
level specifying the camera digitizer offset (in A/D counts). If the
EM gain checkbox is not selected, a CCD camera is assumed.</p>
</div>
<div id="Sx1.SSx2.p2" class="ltx_para">
<p class="ltx_p">Note that the outcome of setting the EM gain to 1 and unchecking the
EM gain checkbox is different as the <a href="estimators/ui/LocalizationUncertainty.html" title="" class="ltx_ref">localization uncertainty</a>
of molecular positions is computed using different formulas for CCD
and EMCCD cameras. Also a stochastic nature of EM gain is modeled
in the <a href="DataGeneratorPlugIn.html" title="" class="ltx_ref">generator of simulated data</a>.</p>
</div>
</div>
</div>
<div id="bib" class="ltx_bibliography">
<h1 class="ltx_title ltx_title_bibliography">References</h1>

<ul id="L1" class="ltx_biblist">
<li id="bib.bib27" class="ltx_bibitem ltx_bib_inproceedings">
<span class="ltx_bibtag ltx_bib_key ltx_role_refnum">[1]</span>
<span class="ltx_bibblock"><span class="ltx_text ltx_bib_author">J. Janesick, K. Klaasen and T. Elliott</span><span class="ltx_text ltx_bib_year">(1985)</span>
</span>
<span class="ltx_bibblock"><span class="ltx_text ltx_bib_title">CCD charge collection efficiency and the photon transfer technique</span>,
</span>
<span class="ltx_bibblock"><span class="ltx_text ltx_bib_pages"> pp. 7–19</span>.
</span>
<span class="ltx_bibblock ltx_bib_cited">Cited by: <a href="#Sx1.p1" title="Camera setup" class="ltx_ref"><span class="ltx_text ltx_ref_title">Camera setup</span></a>.
</span>
</li>
<li id="bib.bib37" class="ltx_bibitem ltx_bib_article">
<span class="ltx_bibtag ltx_bib_key ltx_role_refnum">[2]</span>
<span class="ltx_bibblock"><span class="ltx_text ltx_bib_author">L. J. Van Vliet, F. R. Boddeke, D. Sudar and I. T. Young</span><span class="ltx_text ltx_bib_year">(1998)</span>
</span>
<span class="ltx_bibblock"><span class="ltx_text ltx_bib_title">Image detectors for digital image microscopy</span>,
</span>
<span class="ltx_bibblock"><span class="ltx_text ltx_bib_journal">Digital Image Analysis of Microbes: Imaging, Morphometry, Fluorometry, and Motility Techniques and Applications</span>, <span class="ltx_text ltx_bib_pages"> pp. 37–63</span>.
</span>
<span class="ltx_bibblock ltx_bib_cited">Cited by: <a href="#Sx1.p1" title="Camera setup" class="ltx_ref"><span class="ltx_text ltx_ref_title">Camera setup</span></a>.
</span>
</li>
</ul>
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